Scanning electron microscopy in liver biopsy interpretation in children: a mini atlas

We have extensively studied the ultrastructural picture of the liver by scanning electron microscopy ( SEM) but these studies were not used, up to now, in clinical practice because they were considered to be mainly a means of research in the 3D structure of liver specimens. Our new technique allows us to introduce ourselves to the 3D structure of intracellular organelles, making it possible to study them in normal and pathologic conditions. We used a very small part of the liver biopsies from 5 children aged 3 to 8 years old, who underwent a liver biopsy for diagnostic purposes. The specimens were fixed and processed according to our modification of the OsO 4 maceration method of Tanaka and Mitsushima. Liver biopsies fixed for 20’ in a mixture of glutaraldehyde and paraformaldehyde, postfixed in 1% OsO 4 for 2 h, cut with a tissue sectioner and then macerated in 0.1% OsO 4 for 60 h at room temperature. Specimens were dehydrated in graded acetone, critical point dried and coated with gold palladium. To selectively remove cell components, some specimens were subjected to ultrasound treatment (25 Hz for 1’) prior to dehydration. To demonstrate the hepatic stroma, some aldehyde-fixed specimens were submitted to maceration with NaOH 1N according to Ohtani method. With this method, all cells were removed, allowing the visualization of collagen fibers. Observation was carried out by an Hitachi S4000 Field Emission SEM (Hitachi High-Technologies Co., Tokyo, Japan) operated at 20 kV. We are showing the results of our new technique applied to the liver tissue. These data open, in our opinion, a new field in the research of nuclear pathology, with possible intriguing data on pathological nuclear pore changes in the setting of different liver diseases.